*Target (protein/gene name): FlaA Flagellar outer sheath protein
*NCBI Gene #: Cj1339c *Protein ID:2GNA *Organism (including strain):Leptospira Interrogans Etiologic Risk Group (see link below):BSL1 */Disease Information (sort of like the Intro to your Mini Research Write up): Leptospira interrogans is a thin, helical shaped bacterium that causes the disease Leptospirosis. Leptospirosis is one of the most wide spread zoonotic diseases, and can be deadly when left untreated. The bacterium are able to survive in soil and water for prolonged periods of time until they find a sufficient host. The disease is spread from animals to humans through bodily fluids, such as saliva or contaminated water. This poses a substantial threat in developing countries that lack clean drinking water.
Link to TDR Targets page (if present):N/A Link to Gene Database page:https://www.ncbi.nlm.nih.gov/gene/905631 Essentiality of this protein:FlaA is a protein present on Leptospira interrogans. FlaA is a key component in leptospiral transitional motility, cellular shape, and virulence. Thus, without FlaA the Leptospira interrogans cannot move. Due to their lack of mobility, the bacteria can not spread when FlaA is inhibited. https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3370569/ http://iai.asm.org/content/80/6/2019.figures-only Is it a monomer or multimer as biological unit? Dimer Complex of proteins?:2 Chains Druggable Target:https://www.ncbi.nlm.nih.gov/pubmed/?term=16651261 *EC#:4.2.1.115 Link to BRENDA EC# page:http://www.brenda-enzymes.org/enzyme.php?ecno=4.2.1.115 Enzyme Assay information (spectrophotometric, coupled assay ?, reagents): Dark field microscopy in conjunction with mid log cultures of leptospires can be used in order to observe motility. This requires an Olympus BX-53 139 microscope connected to a Hamamatsu Orca Flash 2.8 camera. Images are taken with CellSense Software, and videos were taken with MicroManager 1.4. The motility was also evaluated in triplicate onto 0.5% agar EMJH plates. LB Agar Plate: $1.50 per plate Olympus BX-53: Must request quote per inquiry, cost is likely >$200 Orca Flash 2.8: Must request quote per inquiry, cost is likely >$200 CellSense: Part of Olympus BX-53 MicroManager: Free https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4820103/ Structure (PDB or Homology model) -- PDB # or closest PDB entry if using homology model: 2GNA After running BLASTP with the amino acid sequence displayed above, it appears the closest human protein to flaA1 has a max score of 48.5. The protein is: putative short-chain dehydrogenase/reductase family 42E member 2 isoform X1 [Homo sapiens], and has an accession number of: XP_016879466. Current Inhibitors:CMP-pseudaminic acid Expression Information (has it been expressed in bacterial cells):Helicobacter pylori Purification Method:His tagged protein Image of protein (PyMol with features delineated and shown separately):
*Amino Acid Sequence (paste as text only - not as screenshot or as 'code'): MPNHQNMLDNQTILITGGTGSFGKCFVRKVLDTTNAKKIIVYSRDELKQSEMAMEFNDPR MRFFIGDVRDLERLNYALEGVDICIHAAALKHVPIAEYNPLECIKTNIMGASNVINACLK NAISQVIALSTDKAANPINLYGATKLCSDKLFVSANNFKGSSQTQFSVVRYGNVVGSRGS VVPFFKKLVQNKASEIPITDIRMTRFWITLDEGVSFVLKSLKRMHGGEIFVPKIPSMKMT DLAKALAPNTPTKIIGIRPGEKLHEVMIPKDESHLALEFEDFFIIQPTISFQTPKDYTLT KLHEKGQKVAPDFEYSSHNNNQWLEPDDLLKLL *length of your protein in Amino Acids:333 Molecular Weight of your protein:37.354 kDa Molar Extinction coefficient of your protein at 280 nm wavelength:21680 TMpred graph Image (http://www.ch.embnet.org/software/TMPRED_form.html).
*NCBI Gene #: Cj1339c
*Protein ID: 2GNA
*Organism (including strain): Leptospira Interrogans
Etiologic Risk Group (see link below): BSL1
*/Disease Information (sort of like the Intro to your Mini Research Write up):
Leptospira interrogans is a thin, helical shaped bacterium that causes the disease Leptospirosis. Leptospirosis is one of the most wide spread zoonotic diseases, and can be deadly when left untreated. The bacterium are able to survive in soil and water for prolonged periods of time until they find a sufficient host. The disease is spread from animals to humans through bodily fluids, such as saliva or contaminated water. This poses a substantial threat in developing countries that lack clean drinking water.
Link to TDR Targets page (if present): N/A
Link to Gene Database page: https://www.ncbi.nlm.nih.gov/gene/905631
Essentiality of this protein:FlaA is a protein present on Leptospira interrogans. FlaA is a key component in leptospiral transitional motility, cellular shape, and virulence. Thus, without FlaA the Leptospira interrogans cannot move. Due to their lack of mobility, the bacteria can not spread when FlaA is inhibited.
https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3370569/
http://iai.asm.org/content/80/6/2019.figures-only
Is it a monomer or multimer as biological unit? Dimer
Complex of proteins?: 2 Chains
Druggable Target: https://www.ncbi.nlm.nih.gov/pubmed/?term=16651261
*EC#:4.2.1.115
Link to BRENDA EC# page: http://www.brenda-enzymes.org/enzyme.php?ecno=4.2.1.115
Enzyme Assay information (spectrophotometric, coupled assay ?, reagents):
Dark field microscopy in conjunction with mid log cultures of leptospires can be used in order to observe motility. This requires an Olympus BX-53 139 microscope connected to a Hamamatsu Orca Flash 2.8 camera. Images are taken with CellSense Software, and videos were taken with MicroManager 1.4. The motility was also evaluated in triplicate onto 0.5% agar EMJH plates.
LB Agar Plate: $1.50 per plate
Olympus BX-53: Must request quote per inquiry, cost is likely >$200
Orca Flash 2.8: Must request quote per inquiry, cost is likely >$200
CellSense: Part of Olympus BX-53
MicroManager: Free
https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4820103/
Structure (PDB or Homology model)
-- PDB # or closest PDB entry if using homology model: 2GNA
After running BLASTP with the amino acid sequence displayed above, it appears the closest human protein to flaA1 has a max score of 48.5. The protein is: putative short-chain dehydrogenase/reductase family 42E member 2 isoform X1 [Homo sapiens], and has an accession number of: XP_016879466.
Current Inhibitors: CMP-pseudaminic acid
Expression Information (has it been expressed in bacterial cells): Helicobacter pylori
Purification Method: His tagged protein
Image of protein (PyMol with features delineated and shown separately):
*Amino Acid Sequence (paste as text only - not as screenshot or as 'code'): MPNHQNMLDNQTILITGGTGSFGKCFVRKVLDTTNAKKIIVYSRDELKQSEMAMEFNDPR
MRFFIGDVRDLERLNYALEGVDICIHAAALKHVPIAEYNPLECIKTNIMGASNVINACLK
NAISQVIALSTDKAANPINLYGATKLCSDKLFVSANNFKGSSQTQFSVVRYGNVVGSRGS
VVPFFKKLVQNKASEIPITDIRMTRFWITLDEGVSFVLKSLKRMHGGEIFVPKIPSMKMT
DLAKALAPNTPTKIIGIRPGEKLHEVMIPKDESHLALEFEDFFIIQPTISFQTPKDYTLT
KLHEKGQKVAPDFEYSSHNNNQWLEPDDLLKLL
*length of your protein in Amino Acids: 333
Molecular Weight of your protein: 37.354 kDa
Molar Extinction coefficient of your protein at 280 nm wavelength: 21680
TMpred graph Image (http://www.ch.embnet.org/software/TMPRED_form.html).